1. Field of the Invention
The present invention relates generally to the fields of molecular biology and medicine. More particularly, it concerns methods and compositions for the production of hematopoietic progenitor cells from embryonic stem cells.
2. Description of Related Art
Due to the significant medical potential of hematopoietic stem and progenitor cells, substantial work has been done to try to improve methods for the differentiation of hematopoietic progenitor cells from pluripotent stem cells. In the human adult, a small number of hematopoietic stem cells present primarily in bone marrow produce heterogeneous populations of actively dividing hematopoietic (CD34+) progenitor cells that differentiate into all the cells of the blood system. However, the CD34+ marker is an imprecise definition of hematopoietic cells since other cell types, notably endothelial cells (blood vessels), also express CD34. Other markers, such as the CD43 marker, may also be used to help identify hematopoietic progenitor cells (e.g., Kadaja-Saarepuu et al., 2007; Vodyanik et al., 2006). In an adult human, hematopoietic progenitors proliferate and differentiate resulting in the generation of hundreds of billions of mature blood cells daily. Hematopoietic progenitor cells are also present in cord blood. In vitro, human embryonic stem cells are capable of indefinite proliferation in culture and are thus capable, at least in principle, of supplying cells and tissues for the replacement of failing or defective human tissue.
The culture of human pluripotent cells with feeder cell lines such as mouse fibroblasts, presents the risk of unexpected transformations that have previously been associated with interspecies exposure during co-culture. Since one of the objectives of human pluripotent stem cell cultures is to create tissues which can ultimately be transplanted into a human body, it is highly desirable that the stem cells are not exposed to cells of another species or to a medium which has been used to culture cells of another species. Accordingly, defining a culture condition that will permit the differentiation of human pluripotent stem cells into the hematopoietic lineage without a co-culture step of any kind, is of great interest in the continued development of techniques for the production of human hematopoietic progenitor cells from human pluripotent stem cells.
Using serum in differentiation medium can also present certain drawbacks and limitations. Serum, e.g., as shown in Chadwick et al. (2003), is an animal product that presents uncertainties regarding the composition of serum across different batches (e.g., regarding variations in the presence and/or concentration of growth factors, etc.). These uncertainties may also contribute to increased variability in the proportion of hematopoietic cells produced across experiments. Additionally, the use of serum may present substantial regulatory issues during clinical development, further complicating commercialization.
There currently exists a clear need for methods of differentiating pluripotent stem cells into hematopoietic progenitor cells without exposing the cells to material from another animal species. Due to the complexities associated with the maintenance and differentiation of pluripotent cells, it is currently not clear how various pluripotent cells may respond to subsequent exposure to growth factors after maintenance in various defined media, as compared to maintenance on mouse feeder cells. Clearly, there is a need for improved methods for the differentiation of pluripotent cells into hematopoietic precursor cells.